Project Summary/Abstract More than one-third of people with epilepsy have inadequate control of seizures even when taking multiple antiepileptic drugs (AEDs). Uncontrolled tonic-clonic seizures place patients at high risk of sudden unexpected death in epilepsy (SUDEP), a major cause of mortality in patients with drug-resistant epilepsy. The high-fat, very low-carbohydrate ketogenic diets (KDs) help some patients with uncontrolled epilepsy, especially children, but have been described as unpalatable and are often not tolerated by patients, leading to high rates of non-compliance. We have obtained preliminary data from a mouse model of Dravet Syndrome (DS), a devastating childhood-onset epilepsy with refractory seizures and a high incidence of SUDEP, and found seizure-induced death is greatly reduced by supplementing their diet with milk whey (WD). This WD was just as protective as two different KDs, but did not require an increase in ketone bodies. Moreover, our preliminary studies suggest that a WD and KDs increase 5-HT levels and 5-HT release in the brain of mice. This is important, as 5-HT is known to inhibit seizures and also plays a critical role in control of breathing. Defects in the 5-HT system have been linked to both respiratory dysfunction and SUDEP. In this proposed project, we will test the hypotheses that 1) whey prevents seizure-induced death by reducing seizure frequency and/or stimulating postictal breathing via increased brain 5-HT, and 2) whey prevents postictal death by reversing the basic underlying defect in DS mice, which is thought to be a decrease in excitability of GABAergic fast- spiking interneurons. To determine how a WD affects seizures, breathing and neurochemistry, we will use video-EEG/EKG to evaluate the anticonvulsant effects of a WD on spontaneous and hyperthermia-induced seizures, and in vivo microdialysis with high performance liquid chromatography to measure extracellular 5-HT in the amygdala, a region implicated in seizure propagation and postictal apnea. We will perform patch clamp recordings on hippocampal brain slices from WD-fed DS mice to evaluate intrinsic electrophysiological properties of interneurons and pyramidal neurons, and will determine if changes in excitability due to the WD are mediated by increased 5-HT signaling. These studies will provide preclinical data that may guide future clinical trials to evaluate milk whey supplementation as a safe and healthy dietary therapy for drug-resistant epilepsy that is more palatable and effective than the KD. These experiments may also lead to new avenues of treatment targeted at the serotonergic mechanisms engaged by milk whey.